Epidemiological studies indicate a higher risk of myocardiac infarction, arrhythmia, stroke and ischemia in young and middle-aged adults who are "occasional" or "regular" cocaine users. Cocaine- and amphetamine-regulated transcript (CART) peptide is a recently identified peptide, the mRNA of which is up-regulated in certain central neurons following acute administration of cocaine or amphetamine. Subsequent studies including those from our laboratory show that CART peptide is expressed in a wide variety of central and peripheral neurons including the sympathetic preganglionic neurons (SPNs), which in turn receive CART-immunoreactive fibers possibly arising from neurons in the medullary C1 area. The central hypothesis is that CART peptide is excitatory to SPNs and that it is up-regulated by cocaine, resulting in an exaggerated sympathetic response in cocaine users. The goal of this project is to characterize in the rat the biological action of CART at the synapse of SPNs, with particular reference to circulation. First, immunohistochemical and tract-tracing studies will determine the location of CART neurons in the medulla or supramedullary nuclei that project to the SPNs. Second, microprobes coated with CART antiserum will be inserted into the intermediolateral cell column (IML) to assay the release of CART-immunoreactive substances from isolated spinal cords in situ or anesthetized rats in vivo. Third, RT-PCR will be used to establish whether or not CART mRNA is up-regulated in the medulla/spinal cord after acute and chronic injection of cocaine (10-20 mg/kg). Fourth, whole-cell patch recordings will be made from single SPNs in the transverse spinal cord slice and the effects of CART peptide on the electrical activity and synaptic transmission of these neurons will be studied. The mechanism and signal transduction pathways underlying the CART action will be investigated. Lastly, the physiological role of CART peptide, which will be administered by intrathecal route on blood pressure and heart rate will be assessed in urethane-anesthetized rats. A number of studies suggest that the sympathetic nervous system plays a major role in cocaine-induced cardiovascular abnormality. To be able to identify the neurotransmitter(s) and the circuitry that respond to cocaine would be a major step toward a better understanding of the intrinsic mechanisms that underlie cocaine-induced vascular disorders. More importantly, identification of putative transmitters involved in these processes would permit a rational approach to the design of novel therapeutics agents for the management of cocaine-induced cardiovascular disorder.